Dicationic biphenyl benzimidazole derivatives as antiprotozoal agents

Article abstract of DOI:10.1016/j.bmc.2004.07.056

A series of biphenyl benzimidazoles diamidines 6a-i were synthesized from their respective diamidoximes, through the bis-O-acetoxyamidoxime followed by hydrogenation in glacial acetic acid/ethanol in the presence of Pd-C. The target compounds contain hydr

Full text of DOI:10.1016/j.bmc.2004.07.056

Bioorganic& Medicinal Chemistry 12 (2004) 5405–5413
Dicationic biphenyl benzimidazole derivatives as
antiprotozoal agents
Mohamed A. Ismail,a Reto Brun,b Tanja Wenzler,b Farial A. Tanious,a
W. David Wilsona and David W. Boykina,
*
aDepartment of Chemistry and Center for Biotechnology and Drug Design, Georgia State University, Atlanta, GA 30303-3083, USA
bSwiss Tropical Institute, Basel, CH4002, Switzerland
Received 7 April 2004; accepted 23 July 2004
Available online 23 August 2004
Abstract—A series of biphenyl benzimidazoles diamidines 6a–i were synthesized from their respective diamidoximes, through the
bis-O-acetoxyamidoxime followed by hydrogenation in glacial acetic acid/ethanol in the presence of Pd–C. The target compounds
contain hydroxy and/or methoxy substituted 1,3-phenyl groups as the central spacer between the two amidino bearing aryl groups.
All of the diamidines showed strong DNA affinities as judged by high DTm values with poly(dAÆdT)2, which varied with structure
and is discussed. Seven of the nine new diamidines gave in vitro IC50 values of approximately 30nM or less versus Trypanosoma
brucei rhodesiense (T.b.r.). Generally the diamidines were less active versus Plasmodium falciparum (P.f.), however one compound
exhibited excellent activity with an IC50 value of 2.1nM. Five of the nine diamidines exhibited excellent in vivo activity in the try-
panosomal STIB900 mouse model giving 3/4 or 4/4 cures at dosage of 20mg/kg ip and three showed similar efficacy at dosage of
10mg/kg or lower.
ꢀ 2004 Elsevier Ltd. All rights reserved.
1. Introduction
which the phenyl group(s) of furamidine have been
replaced with pyridyl group(s) (IIb). Several of these
aza-analogues show in vivo activity, which is superior
to that of furamidine.12 Benzimidazole units are often
key structural elements in the aromatic frame work for
the more effective diamidines.13–16 The benzimidazole
analogue III has been found to bind to DNA in a unique
stacked dimer array, which has potential for develop-
ment of new gene regulation molecules.17–20 Compound
III has shown some activity in an immunosupressed rat
model for Pneumocystis carinii pneumonia17 and in the
STIB900 mouse model for acute stage African trypanoso-
miasis (this manuscript) (Fig. 1).
It has been well documented that aromatic diamidines
exhibit broad-spectrum antimicrobial activity including
effectiveness against the protozoan diseases caused by
Trypanosoma sp. and Plasmodium sp.1 Despite the
broad activity of the aromatic diamidines, pentamidine
(I) is the only compound of this class to be used exten-
sively in the clinic.1 A prodrug, 2,5-bis[4-(methoxyami-
dino)phenyl]furan, of furamidine, (IIa), is an effective
antiprotozoan compound, which is currently entered
into two different Phase II clinical trials as an oral drug
versus human African trypanosomiasis and malaria.1
Many of the active aromatic diamidines for various
structural classes have been shown to bind to the minor
groove of DNA at AT rich sites.2–8 It has been hypoth-
esized that the minor groove binding of these type com-
pounds leads to inhibition of one or more DNA
dependant enzymes, which gives rise to the anti-micro-
bial effect.9–11 Recently, we have made compounds in
Given the promising properties of the various benzimid-
azole aromaticdiamidines and the unusual DNA bind-
ing properties of III we decided to study the effect of
replacing the central furan ring with a 1,3-phenyl group
and we have included hydroxy and/or methoxy substi-
tuted analogues to aid in water solubility. In this work
we also replaced the terminal phenyl group with a pyr-
idyl group. Such alterations in structure offer the poten-
tial to change the base pair recognition on DNA binding
and to yield different absorption and distribution
profiles. We report the synthesis of novel diamidino
Keywords: Diamidines; Benzimidazoles; 1,3-Phenylene; Antiprotozoal
agents; Suzuki coupling.
*
Corresponding author. Tel.: +1 404 651 3798; fax: +1 404 651 1416;
e-mail addresses: dboykin@langate.gsu.edu; dboykin@gsu.edu
0968-0896/$ - see front matter ꢀ 2004 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2004.07.056

5406
M. A. Ismail et al. / Bioorg. Med. Chem. 12 (2004) 5405–5413
O
O
NH
O
HN
NH
HN
X
NH2
H2N
NH2
NH2
IIa; X = CH
IIb; X = N
I
N
O
HN
H2N
HN
NH
III
H2N
Figure 1.
biphenyl benzimidazole derivatives and their evaluation
versus Trypanosoma b. rhodesiense (T.b.r.) and Plasm-
odium falciparum (P.f.).
of an ortho hydroxy group does not cause a reduction
in DTm values as can be seen from the values for 6a
and 6d. This may be due to the smaller size of the hydr-
oxyl group and/or intramolecular hydrogen bonding be-
tween it and a benzimidazole nitrogen atom. The multi-
substituted hydroxyl analogue 6c displays a somewhat
lowered DTm but, this may be a consequence of the but-
tressing effect of the methoxy group yielding greater tor-
sion angle twist. The presence of a meta-amidine unit
(6b) lowers the DTm value relative to its para isomer.
This effect has been noted previously in other systems
and has been attributed to a reduction in complementar-
ity of the curvature of the diamidine compared to the
minor groove.21 Replacement of the terminal phenyl
group with a pyridyl one does not appreciably alter
the DNA affinity (cf. the values for 6f with 6i and that
of 6e with 6h). Consistent with previous studies, there
2. Results and discussion
2.1. Chemistry
Acetate salts of a series of benzimidazole aromatic
diamidines 6a–i were synthesized from their respective
diamidoximes 5a–i, through the corresponding bis-O-
acetoxyamidoximes followed by hydrogenation in gla-
cial acetic acid/ethanol in the presence of Pd–C (Scheme
1). Compounds 5a–i were obtained in three steps start-
ing with the Suzuki coupling reaction of the appropriate
bromobenzaldehyde 1a–c or haloaryl/hetaryl carbonit-
riles 2a–e with cyanophenylboronic acids or substituted
3-formylphenylboronic acids to form the anticipated 3-
formylbiphenyl carbonitrile analogues 3a–i. Subsequent
condensation of the formyl derivatives 3a–i with 3,4-
diaminobenzonitrile in the presence of equimolar ratio
of 1,4-benzoquinone gave the desired dinitriles 4a–i,
which were allowed to react at room temperature for
24h with a mixture of hydroxylamine hydrochloride
and potassium tert-butoxide in DMSO solution to fur-
nish 5a–i in excellent yields. The hydrochloride salt of
the amidoxime 5a was made by passing hydrogen chlo-
ride gas into ethanolicsolution of its free base.
is no direct correlation between DNA affinity and anti-
1
parasiticatcivity.
The strong affinities and excellent
activities observed are consistent with the hypothesis
that a threshold level of DNA affinity is required for
1
antiparasiticactivity.
The new diamidines, except the meta analogue 6b, are
more effective in vitro versus T.b.r. and all except 6c,
6d and 6h are more effective against P.f. than the proto-
type molecule III. Consequently, replacement of the
furan ring of III by a 1,3-phenyl is clearly advantageous
for increased activity of the benzimidazole types versus
these two protozoan organisms. All of the diamidines
except 6b and 6e exhibit anti-trypanosomal IC50 values
of approximately 30nM or less. Four (6d, 6f, 6h, and
6i) gave IC50 values of approximately 20nM or less.
These benzimidazole diamidines were generally not as
active in vitro against P.f., however 6f shows good activ-
ity with an IC50 value of 27.5nM and 6a shows excellent
activity with an IC50 value of 2.1nM. In general, there is
good selectivity for the parasitic organisms by these new
diamidines as judged from their cellular cytotoxicity
(Table 1). The selectivity indices range from over 100
to over 1000, except for compound 6h.
2.2. Biology
Table 1 contains the results of DNA binding studies and
in vitro evaluation against Trypanosoma b. rhodesiense
(T.b.r.) and Plasmodium falciparum (P.f.) for the diami-
dino biphenyl benzimidazoles. The DNA affinities for
the biphenyl benzimidazoles as reflected by DTm values
for poly(dAÆdT)2 are shown in Table 1. Replacement
of the furan ring of III by a 1,3-phenylene unit does
not significantly alter the DNA affinity (cf. values for
III and 6f). However, introduction of a methyl or a
methoxy group ortho to the triaryl ring junctions (6e,
6g, 6h) results in lowering of the DTm values. This reduc-
tion in affinity is presumably due to torsion angle rota-
tion of the triaryl system, which would result in poorer
van der Waals interactions between the aryl rings and
the walls of the minor groove. In contrast, substitution
The results for the in vivo evaluation of the diamidines
versus T.b.r. are shown in Table 2. Five (6c, 6d, 6f, 6h,
and 6i) of the eight diamidines tested in vivo showed
excellent activity curing three or more of the four ani-
mals used in each test at an ip dosage of 20mg/kg or less.

M. A. Ismail et al. / Bioorg. Med. Chem. 12 (2004) 5405–5413
5407
R4
Y
X
NC
(ii)
R3
2a-e
R3
R2
R2
(iii)
(i)
R4
CHO
Br
CHO
X
R1
R1
NC
3a-i
1a-c
R3
R3
R2
R2
(iv)
R4
R4
N
N
X
R1 HN
HN
X
X
R1
HON
NC
NH2
4a-i
5a-i
NOH
CN
H2N
R1
R2
R3
R4
CN
p-CN
3,4
a;
(v)
CH
CH
CH
H
H
H
OH
OH
OH
H
H
H
H
b;
c;
m-CN
p-CN
p-CN
H
R3
OMe
R2
R4
d;
e;
CH
OH
H
H
H
N
OMe
H
p-CN
p-CN
CH
CH
H
H
H
H
H
H
X
R1 HN
HN
f;
NH2
NH
g;
Me p-CN
6a-i
CH
N
H
H
H
H
H
H
H
H2N
h;
i;
OMe
H
p-CN
p-CN
H
H
N
Scheme 1. Reagents and conditions: (i) 3- or 4-cyanophenyl boronic acid, Pd(PPh3)4; (ii) substituted 3-formyl phenyl boronicacid, Pd(PPh 3)4; (iii)
3,4-diaminobenzonitrile, 1,4-benzoquinone, EtOH, reflux; (iv) NH2OHÆHCl/KO-t-Bu, DMSO; (v) (a) AcOH/Ac2O, (b) H2/Pd–C, AcOH.
Table 1. DNA affinities and in vitro anti-protozoan data for biphenyl benzimidazole analogues
R3
R2
R4
N
HN
R1
X
A
A
Code
X
R1
R2
R3
R4
A
DTm
IC50 T.b.r.a,b nM
IC50 P.f. a,b nM
TD50 L6Cellsc lM
I
NA
CH
CH
CH
CH
CH
CH
CH
CH
CH
N
NA
NA
NA
H
NA
NA
NA
OH
OH
OH
H
NA
NA
NA
H
NA
NA
NA
H
p-(C@NH)NH2
p-(C@NH)NH2
p-(C@NH)NH2
p-(C@NH)NH2
m-(C@NH)NH2
p-(C@NH)NH2
p-(C@NH)NH2
p-(C@NH)NH2
p-(C@NH)NH2
p-(C@NH)NH2
p-(C@NH)NH2
p-(C@NH)NH2
12.6
25
2.8
4.5
122
27
64.2
15.5
96
11.4
6.4
IIa
III
6a
6b
6c
6d
6e
6f
24.6
25.3
18.9
20.4
25.3
13.7
25.6
17.1
13.1
24.4
10.1
31.7
117
19.4
37
2.1
H
H
H
214
31
34.3
131
131
40
H
OMe
H
H
OH
H
H
17
54
OMe
H
H
H
19.3
23
H
H
H
4.4
23
27.5
65
6g
6h
6i
H
H
H
Me
H
13.5
23.3
117
H
OMe
H
H
14
15
364
96.4
N
H
H
H
a The T.b.r. strain used was STIB 900 and the P.f. strain was K1; See Refs. 12 and 22 for details.
b Average of duplicate determinations. DNA: polydAÆpolydT; buffer: MES10; Ratio (compound/DNA): 0.3.
c Cytotoxicity was evaluated using cultured L-6 rat myoblast cells using an Alamar Blue assay.

5408
M. A. Ismail et al. / Bioorg. Med. Chem. 12 (2004) 5405–5413
3.2.1. 30-Formyl-40-hydroxybiphenyl-4-carbonitrile (3a).
To a stirred solution of 5-bromosalicyaldehyde (804mg,
4mmol), and tetrakis(triphenylphosphine) palladium
(230mg) in toluene (8mL) under a nitrogen atmosphere
was added 4mL of a 2M aqueous solution of Na2CO3
followed by 4-cyanophenylboronic acid (657mg,
4.8mmol) in 4mL of methanol. The vigorously stirred
mixture was warmed to 80ꢁC for 12h. The solvent was
evaporated, the precipitate was partitioned between
methylene chloride (150mL) and 2M aqueous Na2CO3
(12mL) containing 2mL of concentrated ammonia.
The organiclayer was dried (Na 2SO4), and then concen-
trated to dryness under reduced pressure to afford 3a in
62% yield; mp 143.5–144ꢁC (EtOH). 1H NMR (DMSO-
d6); d 7.13 (d, J = 8.7Hz, 1H), 7.83–7.91 (m, 4H), 7.94
(d, J = 8.7Hz, 1H), 8.02 (s, 1H), 10.30 (s, 1H) 11.00
(br s, 1H). 13C NMR (DMSO-d6); d 191.0, 161.1,
143.3, 134.7, 132.8, 129.3, 127.3, 126.8, 122.6, 118.8,
118.2, 109.5. MS (m/z, rel int.); 223 (M+, 100), 204
(10), 177 (15), 164 (10), 140 (15). HRMS calcd for
C14H9NO2 ms 223.06333. Observed 223.06219. Anal.
(C14H9NO2) C, H, N.
Table 2. In vivo anti-trypanosomal activity of biphenyl benzimidazole
analogues in the STIB900 mouse modela,b
Compound
Dosagec (mg/kg) Curesd Survival (days)e
Pentamidine 20
Furamidine (IIa) 20
0/4
0/4
1/4
0/4
3/4
4/4
2/4
2/4
4/4
3/4
2/4
3/4
3/4
4/4
4/4
42.75
52.5
III
6a
6c
6d
20
20
20
20
5
>27.75
35.25
>58
>60
>46.5
>56.5
>60
6e
6f
20
20
5
>49
6g
6h
20
10
5
>47.5
>53
>60
6i
20
10
>60
>60
a See Ref. 12 for details of STIB900 model.
b IC50 value for 6b did not meet criteria for entry into animal studies.
c Dosage was intraperitoneal.
d Number of mice that survive and are parasite free for 60days.
e Average days of survival; untreated control animals expire between
day 7 and 8 post infection.
3.2.2. 2-(40-Cyano-4-hydroxybiphenyl-3-yl)-1H-benzimid-
azole-5-carbonitrile (4a). A solution of 3a (557.5mg,
Three of the dications (6f, 6h, and 6i) provided cures of
three of four or four of four mice at lower dosage (5 or
10mg/kg). These compounds show significant improve-
ment of in vivo efficacy over the prototype dications
furamidine (IIa) and III. Clearly, these compounds
merit further evaluation in other animal models.
2.5mmol),
3,4-diaminobenzonitrile
(332.5mg,
2.5mmol), and 1,4-benzoquinone (270.2mg, 2.5mmol)
in ethanol (40mL) was allowed to reflux under nitrogen
for overnight. The reaction mixture was distilled off
under reduced pressure. The residue was triturated with
ether and filtered off to afford 4a in 90% yield, mp
>340ꢁC (EtOH). 1H NMR (DMSO-d6); d 7.20 (d,
J = 8.4Hz, 1H), 7.68 (d, J = 8.4Hz, 1H), 7.86–7.98 (m,
6H), 8.28 (s, 1H), 8.57 (s, 1H), 12.80 (br s, 1H), 13.65
(br s, 1H). 13C NMR (DMSO-d6); d 158.4, 153.9
143.5, 132.9, 131.0, 129.3, 126.7, 125.7, 119.7, 118.9,
118.1, 112.8, 109.4, 104.5. MS (m/z, rel int.); 336 (M+,
100), 307 (25), 280 (5), 164 (10). HRMS calcd for
C21H12N4O ms 336.10111. Observed 336.10189. Anal.
(C21H12N4OÆ0.25H2O) C, H, N.
3. Experimental section
3.1. Biology
In vitro assays with Trypanosoma b. rhodesiense STIB
900 and Plasmodium falciparum K1 strain as well as effi-
cacy study in an acute mouse model for T.b. rhodesiense
STIB 900 were carried out as described elsewhere.12,22
3.2.3. 2-[4-Hydroxy-40-(N-hydroxyamidino)-biphenyl-3-
yl]-1H-benzimidazole-5-N-hydroxyamidine hydrochloride
salt (5a). A mixture of hydroxylamine hydrochloride
(1.04g, 15mmol, 10equiv) in anhydrous DMSO (8mL)
was cooled to 5ꢁC under nitrogen and potassium t-but-
oxide (1.68g, 15mmol, 10equiv) was added in portions.
The mixture was stirred for 30min. This mixture was
added to the bis cyanoderivative 4a (1.5mmol, 1equiv).
The reaction mixture was stirred overnight at room tem-
perature. The reaction mixture was then poured slowly
onto ice-water (100mL). The precipitate was filtered
and washed with water to afford 5a (free base) in 94%
3.2. Chemistry
Melting points were recorded using a Thomas–Hoover
(Uni-Melt) capillary melting point apparatus and are
uncorrected. TLC analysis was carried out on silica gel
60 F254 precoated aluminum sheets and detected under
1
UV light. H and 13C NMR spectra were recorded for
all compounds expect 6a and 6h employing a Varian
Unity Plus 300 spectrometer, a Varian GX400 was used
for 6a and 6h. Chemical shifts (d) are in ppm relative to
TMS as internal standard. Mass spectra were recorded
on a VG analytical 70-SE spectrometer (ion source EI,
unless otherwise stated). Elemental analyses were ob-
tained from Atlantic Microlab Inc. (Norcross, GA)
and are within 0.4 of the theoretical values (Table 3).
The compounds reported as salts frequently analyzed
correctly for fractional moles of water and/or ethanol
of solvation. In each case proton NMR showed the pres-
ence of indicated solvent(s). All chemicals and solvents
were purchased from Aldrich Chemical Co., Fisher Sci-
entific, Frontier, or Lancaster.
1
yield; mp 319–322ꢁC. H NMR (DMSO-d6); d 5.87 (s,
4H), 7.13 (d, J = 8.4Hz, 1H), 7.63 (d, J = 8.4Hz, 1H),
7.69–8.02 (m, 7H), 8.45 (s, 1H), 9.60 (s, 1H), 9.63 (s,
1H), 13.20 (br s, 1H), 13.41 (br s, 1H). (5a hydrochoride
salt), mp 301–303ꢁC. Anal. (C21H18N6O3Æ3.0HClÆ2.8-
H2O) C, H, N.
3.2.4. 2-(40-Amidino-4-hydroxybiphenyl-3-yl)-1H-benz-
imidazole-5-amidine acetate salt (6a). To a solution of
5a (402mg, 1mmol) in glacial acetic acid (10mL) was

M. A. Ismail et al. / Bioorg. Med. Chem. 12 (2004) 5405–5413
5409
Table 3. Elemental analysis data
Compound
Calculated
Found
H
C
H
N
C
N
3a (C14H9NO2)
75.32
73.99
44.86
55.62
75.32
73.80
60.64
57.03
71.13
72.12
59.85
54.38
75.32
74.98
58.73
57.55
75.94
75.42
59.34
56.65
81.14
62.36
56.83
81.42
76.95
62.87
57.35
71.78
57.33
54.56
74.98
74.75
55.54
56.78
4.06
3.69
4.76
5.79
4.06
3.71
4.72
5.67
4.37
3.85
4.79
5.89
4.06
3.59
4.92
5.91
4.67
4.03
5.25
6.00
4.37
4.98
6.00
5.01
4.40
5.32
6.24
3.72
4.92
5.79
3.87
3.45
5.12
5.62
6.27
75.00
73.97
44.66
55.30
75.17
73.46
60.60
56.82
70.91
72.30
59.59
54.00
75.21
74.63
58.60
57.44
75.81
75.50
59.08
56.30
80.84
62.03
56.60
81.30
76.75
62.59
57.08
71.51
57.12
54.78
74.80
74.48
55.75
56.96
4.28
3.66
4.51
5.59
4.01
3.62
4.58
5.50
4.48
3.67
4.93
5.75
4.07
3.68
4.77
5.67
4.70
4.09
5.14
5.76
4.43
5.07
5.78
5.02
4.34
5.10
6.00
3.72
4.93
5.90
3.90
3.67
5.14
5.64
5.93
4a (C21H12N4OÆ0.25H2O)
5a (C21H18N6O3Æ3.0HClÆ2.8H2O)
6a (C21H18N6OÆ3.0AcOHÆ1.8H2O)
3b (C14H9NO2)
16.43
14.94
14.41
16.48
14.60
14.42
4b (C21H12N4OÆ0.3H2O)
5b (C21H18N6O3Æ0.75H2O)
6b (C21H18N6OÆ3.0AcOHÆ1.0H2O)
3c (C15H11NO3)
14.78
14.99
4c (C22H14N4O2)
5c (C22H20N6O4Æ0.5H2O)
6c (C22H20N6O2Æ3.0AcOHÆ2.1H2O)
3d (C14H9NO2)
13.58
16.65
14.59
13.31
16.52
14.30
4d (C21H12N4O)
5d (C21H18N6O3Æ1.5H2O)
6d (C21H18N6OÆ2.8AcOHÆ0.8H2OÆ0.5EtOH)
3e (C15H11NO2)
4e (C22H14N4O)
5e (C22H20N6O3Æ1.6H2O)
6e (C22H20N6OÆ2.6AcOHÆ2.0H2O)
3f (C14H9NO)
14.58
14.72
14.90
14.75
5f (C21H18N6O2Æ1.0H2O)
6f (C21H18N6Æ3.0AcOHÆ2.0H2O)
3g (C15H11NO)
4g (C22H14N4Æ0.5H2O)
5g (C22H20N6O2Æ1.1H2O)
6g (C22H20N6Æ2.8AcOHÆ2.3H2O)
4h (C21H13N5O)
14.54
19.93
14.80
19.86
5h (C21H19N7O3Æ1.25H2O)
6h (C21H19N7OÆ3.0AcOHÆ1.5H2O)
3i (C13H8N2O)
16.49
16.33
4i (C20H11N5)
5i (C20H17N7O2Æ2.5H2O)
6i (C20H17N7Æ2.8AcOHÆ1.0H2O)
22.67
18.10
22.29
17.87
slowly added acetic anhydride (0.35mL). After stirring
for overnight TLC indicated complete acylation of the
starting material, the solvent was removed under re-
duced pressure, then to the acetoxy derivative in 40mL
glacial acetic acid/ethanol (1:3) was added 10% palla-
dium on carbon (80mg). The mixture was placed on
Parr hydrogenation apparatus at 50psi for 4h at room
temperature. The mixture was filtered through hyflo
and the filter pad washed with water. The filtrate was
evaporated under reduced pressure and the precipitate
was collected and washed with ether to give 6a in
118.0, 112.0. MS (m/z, rel int.); 223 (M+, 100), 204
(10), 193 (5), 177 (20), 166 (15), 140 (15). Anal.
(C14H9NO2) C, H.
3.2.6. 2-(30-Cyano-4-hydroxybiphenyl-3-yl)-1H-benzimid-
azole-5-carbonitrile (4b). The same procedure described
for 4a was used starting with 3b. Yield 89%, mp
>340ꢁC (EtOH). 1H NMR (DMSO-d6);
d 7.20
(d, J = 8.4Hz, 1H), 7.67–7.75 (m, 2H), 7.82–7.88 (m,
3H), 8.10 (d, J = 8.4Hz, 1H), 8.21 (s, 1H), 8.26 (s,
1H), 8.55 (s, 1H), 13.20 (br s, 2H). 13C NMR; d 158.2,
154.0, 140.2, 130.8, 130.7, 130.5, 130.2, 129.4, 129.1,
126.3, 125.4, 119.7, 118.8, 118.0, 112.7, 112.1, 104.7.
MS (m/z, rel int.); 336 (M+, 100), 307 (20), 306 (12),
168 (5), 140 (5). HRMS calcd for C21H12N4O:
336.10111. Observed 336.10247. Anal. (C21H12N4O–
0.3H2O) C, H.
1
78.5% yield, mp 223–224ꢁC. H NMR (D2O/DMSO-
d6); d 1.80 (s, 3 · CH3), 7.00 (d, J = 8.4Hz, 1H), 7.48
(d, J = 8.4Hz, 1H), 7.64–7.67 (m, 2H), 7.90 (s, 4H),
8.08 (s, 1H), 8.62 (s, 1H). Anal. (C21H18N6OÆ3.0Ac-
OHÆ1.8H2O) C, H, N.
3.2.5. 30-Formyl-40-hydroxybiphenyl-3-carbonitrile (3b).
The same procedure described for 3a was used employ-
ing 3-cyanophenylboronic acid instead of 4-cyano-
3.2.7. 2-[4-Hydroxy-30-(N-hydroxyamidino)-biphenyl-3-
yl]-1H-benzimidazole-5-N-hydroxyamidine (5b). The
same procedure described for 5a was used starting with
phenylboroniciadc. Yield 70%; mp 139–140
d
ꢁC
7.12 (d,
(EtOH). 1H NMR (DMSO-d6);
J = 8.7Hz, 1H), 7.62–7.81 (m, 2H), 7.92–8.02 (m,
4b. Yield 97%, mp >340ꢁC. H NMR (DMSO-d6); d
1
5.89 (s, 4H), 7.20 (d, J = 8.4Hz, 1H), 7.58 (d,
J = 8.4Hz, 1H), 7.63–8.90 (m, 6H), 8.09 (s, 1H), 8.50
(s, 1H), 9.70 (s, 2H), 13.20 (br s, 1H), 13.44 (br s, 1H).
13C NMR (DMSO-d6); d 157.7, 152.4, 151.2, 150.9,
3H), 8.14 (s, 1H), 10.32 (s, 1H) 11.00 (br s, 1H). 13C
NMR (DMSO-d6); d 191.3, 160.7, 140.0, 134.6,
130.8, 130.5, 130.1, 129.5, 129.2, 127.4, 122.5, 118.7,

5410
M. A. Ismail et al. / Bioorg. Med. Chem. 12 (2004) 5405–5413
139.3, 134.1, 131.2, 130.2, 128.7, 126.7, 124.5, 124.1,
123.4, 117.8, 115.0, 112.8, 108.6. Anal.
(C21H18N6O3Æ0.75H2O) C, H.
NMR (CDCl3); d 7.25 (t, J = 7.8Hz, 1H), 7.60–7.70
(m, 2H), 7.73–7.80 (m, 4H), 9.97 (s, 1H), 11.64 (s, 1H).
13C NMR (DMSO-d6); d 197.4. 157.5, 141.0, 137.4,
134.2, 132.1, 130.0, 128.0, 121.5, 120.4, 118.8, 110.0.
MS (m/z, rel int.); 223 (M+, 100), 204 (20), 195 (25),
177 (25), 140 (20). HRMS calcd for C14H9NO2:
223.06333. Observed 223.06256. Anal. (C14H9NO2)
C, H.
3.2.8. 2-(30-Amidino-4-hydroxybiphenyl-3-yl)-1H-benz-
imidazole-5-amidine acetate salt (6b). The same proce-
dure described for 6a was used starting with 5b. Yield
80%, mp 208–209ꢁC. 1H NMR (D2O/DMSO-d6); d
1.80 (s, 3 · CH3), 7.02 (d, J = 8.4Hz, 1H), 7.47 (d,
J = 8.4Hz, 1H), 7.62–7.72 (m, 4H), 8.00 (d, J = 7.2Hz,
1H), 8.07 (s, 1H), 8.11 (s, 1H), 8.66 (s, 1H). Anal.
(C21H18N6OÆ3.0AcOHÆ1.0H2O) C, H, N.
3.2.14. 2-(40-Cyano-2-hydroxybiphenyl-3-yl)-1H-benzim-
idazole-5-carbonitrile (4d). The same procedure de-
scribed for 4a was used starting with 3d. Yield 84%,
mp 318–320ꢁC. 1H NMR (DMSO-d6); d 7.18 (t,
J = 7.5Hz, 1H), 7.56 (d, J = 7.5Hz, 1H), 7.71 (d, J =
8.4Hz, 1H), 7.78 (d, J = 7.5Hz, 1H), 7.84 (d,
J = 8.1Hz, 2H), 7.91 (d, J = 8.1Hz, 2H), 8.14 (d,
J = 8.4Hz, 1H), 8.28 (s, 1H), 13.70 (br s, 1H), 13.90
(br s, 1H). MS (m/z, rel int.); 337 (M+ + 1, 68), 309
(100), 293 (40). Anal. (C21H12N4O) C, H, N.
3.2.9. 50-Formyl-40-hydroxy-30-methoxybiphenyl-4-carbo-
nitrile (3c). The same procedure described for 3a was
used employing 5-bromo-2-hydroxy-3-methoxybenzal-
dehyde instead of 5-bromosalicyaldehyde. Yield 57%;
mp 177–177.5ꢁC. 1H NMR (DMSO-d6); d 3.97 (s,
3H), 7.61 (s, 2H), 7.90 (s, 4H), 10.33 (s, 1H), 10.57 (br
s, 1H). 13C NMR (DMSO-d6); d 191.3, 151.2, 149.0,
143.7, 132.7, 129.2, 127.1, 122.6, 118.9, 118.3, 115.7,
109.5, 56.3. MS (m/z, rel int.); 253 (M+, 100), 210 (8),
207 (10), 177 (10), 154 (15), 127 (18). HRMS calcd for
C15H11NO3: 253.07389. Observed 253.07181. Anal.
(C15H11NO3) C, H.
3.2.15. 2-[2-Hydroxy-40-(N-hydroxyamidino)-biphenyl-3-
yl]-1H-benzimidazole-5-N-hydroxyamidine (5d). The
same procedure described for 5a was used starting with
1
4d. Yield 100%, mp 322–325ꢁC. H NMR (DMSO-d6);
d 6.00 (s, 4H), 7.13 (t, J = 7.8Hz, 1H), 7.49 (d,
J = 7.2Hz, 1H), 7.61–7.77 (m, 6H), 7.88–8.08 (m, 2H),
9.70 (s, 2H), 13.40 (br s, 1H), 13.90 (br s, 1H). Anal.
(C21H18N6O3Æ1.5H2O) C, H.
3.2.10. 2-(40-Cyano-4-hydroxy-5-methoxybiphenyl-3-yl)-
1H-benzimidazole-5-carbonitrile (4c). The same proce-
dure described for 4a was used starting with 3c. Yield
79%, mp 334–335ꢁC. 1H NMR (DMSO-d6); d 3.98
(s, 3H), 7.49 (s, 1H), 7.68 (d, J = 8.4Hz, 1H), 7.84
(d, J = 8.4Hz, 1H), 7.94–8.00 (m, 4H), 8.13 (s,
1H), 8.25 (s, 1H), 13.2 (br s, 2H). 13C NMR; d 154.2,
149.1, 148.9, 143.8, 132.7, 129.0, 126.9, 126.8, 119.6,
118.9, 116.8, 112.8, 112.3, 109.4, 104.7, 99.4, 56.0. MS
(m/z, rel int.); 366 (M+, 100), 348 (42), 337 (20), 323
(30), 307 (10). HRMS calcd for C22H14N4O2:
366.11168. Observed 366.11188. Anal. (C22H14N4O2)
C, H.
3.2.16. 2-(40-Amidino-2-hydroxybiphenyl-3-yl)-1H-benz-
imidazole-5-amidine acetate salt (6d). The same proce-
dure described for 6a was used starting with 5d. Yield
1
90%, mp 228–229.5ꢁC. H NMR (D2O/DMSO-d6); d
1.80 (s, 2.8 · CH3), 6.97 (t, J = 7.8Hz, 1H), 7.35 (d,
J = 7.5Hz, 1H), 7.45 (d, J = 7.5Hz, 1H), 7.60 (d, J =
7.8Hz, 1H), 7.86 (d, J = 8.4Hz, 2H), 7.93 (d,
J = 8.4Hz, 2H), 8.05 (s, 1H), 8.27 (d, J = 7.8Hz, 1H).
MS (m/z, rel int.); 371 (M+ + 1, 5), 354 (5), 337 (100),
336 (50), 307 (10). Anal. (C21H18N6OÆ2.8AcOHÆ0.8-
H2OÆ0.5EtOH) C, H, N.
3.2.11. 2-[4-Hydroxy-40-(N-hydroxyamidino)-5-methoxy-
biphenyl-3-yl]-1H-benzimidazole-5-N-hydroxyamidine (5c).
The same procedure described for 5a was used starting
with 4c. Yield 99%, mp 319–321ꢁC. 1H NMR
(DMSO-d6); d 4.00 (s, 3H), 5.89 (s, 4H), 7.43 (s, 1H),
7.59–7.74 (m, 2H), 7.82–7.88 (m, 4H), 8.05 (s, 1H),
8.09 (s, 1H), 9.61 (s, 1H), 9.69 (s, 1H), 13.40 (br s,
2H). Anal. (C22H20N6O4Æ0.5H2O) C, H.
3.2.17. 30-Formyl-40-methoxybiphenyl-4-carbonitrile (3e).
To a stirred solution of 4-bromobenzonitrile (910mg,
5mmol), and tetrakis(triphenylphosphine) palladium
(288mg) in toluene (10mL) under a nitrogen atmos-
phere was added 5mL of a 2M aqueous solution of
Na2CO3 followed by 3-formyl-4-methoxyphenylboronic
acid (1080mg, 6mmol) in 5mL of methanol. The vigor-
ously stirred mixture was warmed to 80ꢁC for 12h. The
solvent was evaporated, the precipitate was partitioned
between methylene chloride (200mL) and 2M aqueous
Na2CO3 (15mL) containing 3mL of concentrated
3.2.12. 2-(40-Amidino-4-hydroxy-5-methoxybiphenyl-3-
yl)-1H-benzimidazole-5-amidine acetate salt (6c). The
same procedure described for 6a was used starting with
1
5c. Yield 75%, mp 230–231ꢁC. H NMR (D2O/DMSO-
ammonia. The organiclayer was dried (Na SO4), and
2
d6); d 1.80 (s, 3 · CH3), 3.93 (s, 3H), 7.27 (s, 1H), 7.47 (d,
J = 8.4Hz, 1H), 7.63 (d, J = 8.4Hz, 1H), 7.87–7.97 (m,
4H), 8.07 (s, 1H), 8.24 (s, 1H). MS (m/z, rel int.); 383
(M+ꢀNH3, 10), 366 (50), 351 (10), 336 (100). Anal.
(C22H20N6O2Æ3.0AcOHÆ2.1H2O) C, H, N.
then concentrated to dryness under reduced pressure
to afford 3e in 70% yield; mp 146–147ꢁC (SiO2, hexa-
nes/EtOAc, 90:10). 1H NMR (DMSO-d6); d 4.00
(s, 3H), 7.39 (d, J = 9.0Hz, 1H), 7.88–7.94 (m, 4H),
8.03 (d, J = 2.7Hz, 1H), 8.09 (dd, J = 9.0, 2.7Hz, 1H),
10.40 (s, 1H). 13C NMR; d 188.9, 161.7, 143.1, 134.7,
132.8, 130.5, 127.0, 126.2, 124.4, 118.8, 113.7, 109.8,
56.2. MS (m/z, rel int.); 237 (M+, 100), 220 (20), 208
(10), 191 (25), 177 (35), 140 (20). Anal. (C15H11NO2)
C, H.
3.2.13. 30-Formyl-20-hydroxybiphenyl-4-carbonitrile (3d).
The same procedure described for 3a was used employ-
ing 3-bromosalicyaldehyde23 instead of 5-bromosalicyal-
1
dehyde. Yield 58%; mp 120–121ꢁC (hexanes/ether). H

M. A. Ismail et al. / Bioorg. Med. Chem. 12 (2004) 5405–5413
5411
3.2.18. 2-(40-Cyano-4-methoxybiphenyl-3-yl)-1H-benzim-
idazole-5-carbonitrile (4e). The same procedure de-
scribed for 4a was used starting with 3e. Yield 88%,
3.2.23. 2-[40-(N-Hydroxyamidino)biphenyl-3-yl]-1H-benz-
imidazole-5-N-hydroxyamidine (5f). The same procedure
described for 5a was used starting with 4f. Yield 93%,
1
1
mp 289–290ꢁC. H NMR (DMSO-d6); d 4.09 (s, 3H),
mp 317–319ꢁC. H NMR (DMSO-d6); d 5.89 (s, 2H),
7.43 (d, J = 8.4Hz, 1H), 7.61 (d, J = 8.4Hz, 1H), 7.81
(s, 1H), 7.94–8.22 (m, 6H), 8.67 (s, 1H), 12.63 (br s,
1H). 13C NMR (DMSO-d6); d 157.5, 151.1, 143.5,
142.1, 137.9, 132.9, 130.9, 130.7, 128.3, 127.1, 125.7,
125.6, 123.4, 120.1, 118.9, 117.7, 113.3, 113.2, 109.6,
103.8, 56.3. MS (m/z, rel int.); 350 (M+, 100), 321 (40),
306 (12), 144 (50). HRMS calcd for C22H14N4O:
350.11676. Observed 350.11569. Anal. (C22H14N4O) C,
H.
5.92 (s, 2H), 7.60–7.70 (m, 3H), 7.80–7.98 (m, 5H),
8.00 (s, 1H), 8.21 (d, J = 7.8Hz, 1H), 8.52 (s, 1H), 9.60
(s, 1H), 9.74 (s, 1H), 13.11 (br s, 1H). Anal.
(C21H18N6O2Æ1.0H2O) C, H.
3.2.24. 2-(40-Amidinobiphenyl-3-yl)-1H-benzimidazole-5-
amidine acetate salt (6f). The same procedure described
for 6a was used starting with 5f. Yield 72%, mp 211–
212ꢁC. 1H NMR (D2O/DMSO-d6);
d
1.78 (s,
3 · CH3), 7.60 (d, J = 8.4Hz, 1H), 7.69 (t, J = 7.8Hz,
1H), 7.76 (d, J = 8.4Hz, 1H), 7.91 (d, J = 7.8Hz, 1H),
7.96 (d, J = 8.7Hz, 2H), 8.03 (d, J = 8.7Hz, 2H), 8.12
(s, 1H), 8.33 (d, J = 7.8Hz, 1H), 8.65 (s, 1H). Anal.
(C21H18N6Æ3.0AcOHÆ2.0H2O) C, H, N.
3.2.19. 2-[40-(N-Hydroxyamidino)-4-methoxybiphenyl-3-
yl]-1H-benzimidazole-5-N-hydroxyamidine (5e). The
same procedure described for 5a was used starting with
1
4e. Yield 95%, mp >340ꢁC. H NMR (DMSO-d6); d
4.09 (s, 3H), 6.09 (s, 2H), 6.71 (s, 2H), 7.38 (d,
J = 8.4Hz, 1H), 7.58 (d, J = 8.4Hz, 1H), 7.69 (d,
J = 8.4Hz, 1H), 7.75 (d, J = 8.7Hz, 2H), 7.81 (d, J =
8.7Hz, 2H), 7.87 (dd, J = 8.4, 2.4Hz, 1H), 7.99 (s,
1H), 8.64 (d, J = 2.4Hz, 1H), 9.80 (s, 1H), 10.0 (s,
1H), 12.55 (br s, 1H). 13C NMR (DMSO-d6); d 156.6,
151.0, 150.0, 139.7, 137.9, 132.1, 131.5, 129.6, 127.6,
126.1, 125.9, 120.4, 118.0, 112.9, 56.1. FABMS (m/z,
rel int.); 417 (M+ + 1, 100), 401 (58), 394 (30), 368
(20), 350 (10). HRMS calcd for C22H21N6O3:
417.16751. Observed 417.16760. Anal. (C22H20-
N6O3Æ1.6H2O) C, H.
3.2.25. 30-Formyl-2-methylbiphenyl-4-carbonitrile (3g).
The same procedure described for 3f was used employ-
ing 4-bromo-3-methylbenzonitrile instead of 4-bromo-
benzonitrile. Yield 82%, mp 86–86.5ꢁC. 1H NMR
(DMSO-d6); d 2.27 (s, 3H), 7.47 (d, J = 7.8Hz, 1H),
7.71–7.79 (m, 3H), 7.85 (s, 1H), 7.90 (s, 1H), 7.95–7.98
(m, 1H), 10.08 (s, 1H). 13C NMR (DMSO-d6); d
193.0, 144.8, 140.3, 136.8, 136.3, 134.7, 133.9, 130.5,
130.0, 129.8, 129.4, 128.4, 118.7, 110.6, 19.7. Anal.
(C15H11NO) C, H.
3.2.26. 2-(40-Cyano-20-methylbiphenyl-3-yl)-1H-benzimid-
azole-5-carbonitrile (4g). The same procedure described
for 4a was used starting with 3g. Yield 75%, mp 247–
250ꢁC. 1H NMR (DMSO-d6); d 2.26 (s, 3H), 7.50–
7.63 (m, 3H), 7.67–7.80 (m, 3H), 7.87 (s, 1H), 8.17 (d,
J = 7.8Hz, 2H), 8.28 (d, J = 7.8Hz, 1H), 13.40 (br s,
1H). MS (m/z, rel int.); 334 (M+, 100), 215 (5), 190
(20), 167 (6). HRMS calcd for C22H14N4: 334.12185.
Observed 334.12142. Anal. (C22H14N4Æ0.5H2O) C, H.
3.2.20. 2-(40-Amidino-4-methoxybiphenyl-3-yl)-1H-benz-
imidazole-5-amidine acetate salt (6e). The same proce-
dure described for 6a was used starting with 5e. Yield
62%, mp 220–221ꢁC. 1H NMR (D2O/DMSO-d6); d
1.78 (s, 2.6 · CH3), 4.11 (s, 3H), 7.43 (d, J = 8.4Hz,
1H), 7.65 (d, J = 8.4Hz, 1H), 7.81 (d, J = 8.4Hz, 1H),
7.87–7.96 (m, 4H), 8.15 (dd, J = 8.4, 2.4Hz, 1H), 8.31
(s,
1H),
8.69
(d,
J = 2.4Hz,
1H).
Anal.
(C22H20N6OÆ2.6AcOHÆ2.0H2O) C, H, N.
3.2.27. 2-[40-(N-Hydroxyamidino)-20-methylbiphenyl-3-
yl]-1H-benzimidazole-5-N-hydroxyamidine (5g). The
same procedure described for 5a was used starting with
4g. Yield 99%, mp 295–297ꢁC. 1H NMR (DMSO-d6); d
2.30 (s, 3H), 5.87 (s, 4H), 7.32 (d, J = 8.1Hz, 1H), 7.50
(d, J = 7.8Hz, 1H), 7.61–7.97 (m, 6H), 8.15 (s, 1H),
8.21 (d, J = 7.8Hz, 1H), 9.59 (s, 1H), 9.68 (s, 1H),
13.02 (br s, 1H). 13C NMR (DMSO-d6); d 151.5,
150.5, 141.4, 141.0, 134.5, 132.5, 130.3, 130.0, 129.3,
128.9, 127.3, 127.2, 126.7, 125.2, 123.0, 119.8, 118.2,
115.9, 110.7, 108.4, 20.2. FABMS (m/z, rel int.); 401
(M+ + 1, 100), 386 (55), 368 (30), 352 (15), 335 (10).
HRMS calcd for C22H21N6O2: 401.17260. Observed
401.17287. Anal. (C22H20N6O2Æ1.1H2O) C, H.
3.2.21. 30-Formylbiphenyl-4-carbonitrile (3f). The same
procedure described for 3e was used employing 3-form-
ylphenylboroniciadc instead of 3-formyl-4-meth-
1
oxyphenylboronicacid. Yield 80%, mp 122–123 ꢁC. H
NMR (DMSO-d6); d 7.72–7.78 (m, 1H), 7.98–8.03 (m,
5H), 8.11 (m, 1H), 8.29 (s, 1H), 10.13 (s, 1H). 13C
NMR (DMSO-d6); d 193.0, 143.3, 139.0, 136.9, 132.98,
132.95, 130.0, 129.0, 128.5, 127.7, 118.7, 110.6. Anal.
(C14H9NO) C, H.
3.2.22. 2-(40-Cyanobiphenyl-3-yl)-1H-benzimidazole-5-
carbonitrile (4f). The same procedure described for 4a
was used starting with 3f. Yield 81%, mp 303–304ꢁC.
1H NMR (DMSO-d6); d 7.63 (d, J = 8.4Hz, 1H), 7.73
(t, J = 7.8Hz, 1H), 7.80 (d, J = 8.4Hz, 1H), 7.96 (d,
J = 7.8Hz, 1H), 8.00–8.06 (m, 4H), 8.19 (s, 1H), 8.30
(d, J = 7.8Hz, 1H), 8.57 (s, 1H), 13.60 (br s, 1H). 13C
NMR (DMSO-d6); d 153.9, 143.7, 139.0, 132.9, 130.07,
130.01, 129.2, 127.7, 127.1, 125.8, 125.4, 119.9, 118.7,
110.5, 104.1. MS (m/z, rel int.); 320 (M+, 100), 291 (5),
204 (5), 177 (8), 151 (3). HRMS calcd for C21H12N4:
320.10620. Observed 320.10644.
3.2.28. 2-(40-Amidino-20-methylbiphenyl-3-yl)-1H-benz-
imidazole-5-amidine acetate salt (6g). The same proce-
dure described for 6a was used starting with 5g. Yield
83%, mp 201–203ꢁC. 1H NMR (D2O/DMSO-d6); d
1.80 (s, 2.8 · CH3), 2.33 (s, 3H), 7.50–7.59 (m, 2H),
7.62–7.78 (m, 3H), 7.82–7.95 (m, 2H), 8.12 (s, 1H),
8.21–8.34 (m, 2H). Anal. (C22H20N6Æ2.8AcOHÆ2.3H2O)
C, H, N.

5412
M. A. Ismail et al. / Bioorg. Med. Chem. 12 (2004) 5405–5413
3.2.29. 6-(3-Formyl-4-methoxyphenyl)-nicotinonitrile
(3h). The same procedure described for 3e was used
(m/z, rel int.); 208 (M+, 100), 179 (70), 152 (15), 125
(5). HRMS calcd for C13H8N2O: 208.06366. Observed
208.06066. Anal. (C13H8N2O) C, H.
employing 6-chloronicotinonitrile24 instead of 4-bromo-
1
benzonitrile. Yield 66.5%; mp 197–198ꢁC (EtOH). H
NMR (CDCl3); d 4.00 (s, 3H), 7.16 (d, J = 8.4Hz,
1H), 7.87 (d, J = 8.4Hz, 1H), 8.00 (dd, J = 8.4, 2.1Hz,
1H), 8.41 (dd, J = 8.4, 2.1Hz, 1H), 8.46 (d, J =
2.1Hz, 1H), 8.92 (d, J = 2.1Hz, 1H), 10.57 (s, 1H). 13C
NMR (DMSO-d6); d 188.5, 162.7, 157.5, 152.0, 140.4,
134.3, 129.1, 126.7, 124.3, 119.2, 116.8, 113.1, 106.7,
56.1. MS (m/z, rel int.); 238 (M+, 100), 221 (32), 209
(15), 192 (25), 178 (25), 166 (20). HRMS calcd for
C14H10N2O2: 238.07423. Observed 238.07486.
3.2.34. 2-[3-(5-Cyanopyridin-2-yl)-phenyl]-1H-benzimid-
azole-5-carbonitrile (4i). The same procedure described
for 4a was used starting with 3i. Yield 83.5%, mp 281–
282ꢁC. 1H NMR (DMSO-d6); d 7.63 (d, J = 7.8Hz,
1H), 7.76 (t, J = 7.8Hz, 1H), 7.81–7.88 (m, 1H), 8.24–
8.38 (m, 4H), 8.49 (d, J = 7.8Hz, 1H), 9.03 (s, 1H),
9.18 (s, 1H), 13.61 (br s, 1H). 13C NMR (DMSO-d6);
d 158.2, 152.5, 141.1, 137.6, 129.8, 129.2, 128.7, 126.0,
125.5, 124.0, 120.3, 119.9, 117.1, 112.9, 107.8, 104.0.
MS (m/z, rel int.); 321 (M+, 100), 293 (12), 268 (5).
HRMS calcd for C20H11N5: 321.10145. Observed
321.10069. Anal. (C20H11N5) C, H.
3.2.30. 2-[5-(5-Cyanopyridin-2-yl)-2-methoxyphenyl]-1H-
benzimidazole-5-carbonitrile (4h). The same procedure
described for 4a was used starting with 3h. Yield 92%,
mp 316.5–319ꢁC. 1H NMR (DMSO-d6); d 4.07 (s,
3H), 7.44 (d, J = 8.4Hz, 1H), 7.58–7.87 (m, 2H), 8.03–
8.36 (m, 4H), 9.09 (s, 1H), 9.20 (d, J = 2.1Hz, 1H),
12.60 (br s, 1H). 13C NMR (DMSO-d6); d 158.7,
157.9, 152.4, 151.4, 140.8, 130.8, 129.6, 129.1, 125.4,
119.9, 119.4, 117.5, 117.2, 112.9, 106.8, 103.8, 56.3.
MS (m/z, rel int.); 351 (M+, 100), 322 (40), 293 (10),
143 (35). HRMS calcd for C21H13N5O: 351.11201. Ob-
served 351.11067. Anal. (C21H13N5O) C, H, N.
3.2.35. 2-{3-[5-(N-Hydroxyamidino)-pyridin-2-yl]-phen-
yl}-1H-benzimidazole-5-N-hydroxyamidine (5i). The
same procedure described for 5a was used starting with
1
4i. Yield 97%, mp 295–297ꢁC. H NMR (DMSO-d6);
5.82 (s, 2H), 6.08 (s, 2H), 7.51–7.72 (m, 3H), 8.00 (s,
1H), 8.11–8.28 (m, 4H), 8.95 (s, 1H), 9.03 (s, 1H), 9.61
(s, 1H), 9.93 (s, 1H). 13.18 (br s, 1H). Anal.
(C20H17N7O2Æ2.5H2O) C, H, N.
3.2.36. 2-[3-(5-Amidinopyridin-2-yl)-phenyl]-1H-benzimid-
azole-5-amidine acetate salt (6i). The same procedure
described for 6a was used starting with 5i. Yield 73%,
mp 198–200ꢁC. H NMR (D2O/DMSO-d6); d 1.80 (s,
2.8 · CH3), 7.64 (d, J = 8.1Hz, 1H), 7.72–7.81 (m,
2H), 8.14 (s, 1H), 8.24–8.40 (m, 4H), 9.08 (s, 1H), 9.12
(s, 1H). MS (m/z, rel int.); 356 (M+ + 1, 5), 322 (100),
297 (5). Anal. (C20H17N7Æ2.8AcOHÆ1.0H2O) C, H, N.
3.2.31. 2-{5-[5-(N-Hydroxyamidino)-pyridin-2-yl]-2-meth-
oxyphenyl}-1H-benzimidazole-5-N-hydroxyamidine (5h).
The same procedure described for 5a was used starting
with 4h. Yield 99%, mp 276–279ꢁC. 1H NMR
(DMSO-d6); d 4.08 (s, 3H), 5.83 (s, 2H), 6.02 (s, 2H),
7.37 (d, J = 8.4Hz, 1H), 7.58–7.68 (m, 2H), 7.95 (s,
1H), 8.02 (d, J = 8.4Hz, 1H), 8.11 (d, J = 8.4Hz, 1H),
8.22 (dd, J = 8.4, 2.1Hz, 1H), 8.95 (s, 1H), 9.13
(d, J = 2.1Hz, 1H), 9.57 (s, 1H), 9.87 (s, 1H), 12.30 (br
s, 1H). 13C NMR (DMSO-d6); d 157.6, 155.2, 152.1,
151.9, 149.7, 148.8, 146.5, 133.9, 130.9, 129.4, 128.0,
127.2, 118.8, 118.2, 112.6, 56.1. FABMS (m/z, rel int.);
418 (M+ + 1, 70), 401 (40), 385 (23), 327 (50), 237
(100). HRMS calcd for C21H20N7O3: 418.16276. Ob-
served 418.16178. Anal. (C21H19N7O3Æ1.25H2O) C, H.
1
Acknowledgements
This work was supported by NIH Grant GM-61587,
an award from the Bill and Melinda Gates Foundation
and M.A.I. was awarded a Fellowship by ICSC-World
Laboratory.
References and notes
3.2.32. 2-[5-(5-Amidinopyridin-2-yl)-2-methoxyphenyl]-
1H-benzimidazole-5-amidine acetate salt (6h). The same
procedure described for 6a was used starting with 5h.
Yield 80%, mp 231–232ꢁC. 1H NMR (D2O/DMSO-
d6); d 1.83 (s, 3 · CH3), 4.18 (s, 3H), 7.42 (d,
J = 8.4Hz, 1H), 7.63 (d, J = 8.4Hz, 1H), 7.78–7.88 (m,
2H), 8.10–8.33 (m, 3H), 9.00 (s, 1H), 9.17 (s, 1H). MS
(m/z, rel int.); 385 (M+, 4), 351 (100), 337 (75). Anal.
(C21H19N7OÆ3.0AcOHÆ1.5H2O) C, H, N.
1. Tidwell, R. R.; Boykin, D. W. Dicationic DNA Minor
Groove Binders as Antimicrobial Agents; In Small Mole-
cule DNA and RNA Binders: From Synthesis to Nucleic
Acid Complexes; Demeunynck, M., Bailly, C., Wilson, W.
D., Eds.; Wiley-VCH: New York, 2003; Vol. 2, p 414.
2. Czarny, A.; Boykin, D. W.; Wood, A.; Nunn, C. M.;
Neidle, S.; Zhao, M. R. J.; Wilson, W. D. J. Am. Chem.
Soc. 1995, 117, 4716.
3. Laughton, C. A.; Tanious, F. A.; Nunn, C. M.; Boykin, D.
W.; Wilson, W. D.; Neidle, S. Biochemistry 1996, 35, 5655.
4. Trent, J. O.; Clark, G. R.; Kumar, A.; Wilson, D. W.;
Boykin, D. W.; Hall, J. E.; Tidwell, R. R.; Blagburn, B. L.;
Neidle, S. J. Med. Chem. 1996, 39, 4554.
5. Wilson, W. D.; Tanious, F. A.; Ding, D.; Kumar, A.;
Boykin, D. W.; Colson, P.; Houssier, C.; Bailly, C. J. Am.
Chem. Soc. 1998, 120, 10310.
6. Boykin, D. W.; Kumar, A.; Spychala, J.; Zhou, M.;
Lombardy, R.; Wilson, W. D.; Dykstra, C. C.; Jones, S.
K.; Hall, J. E.; Tidwell, R. R.; Laughton, C.; Nunn, C. M.;
Neidle, S. J. Med. Chem. 1995, 38, 912.
3.2.33. 6-(3-Formylphenyl)-nicotinonitrile (3i). The same
procedure described for 3h was used employing 3-form-
ylphenylboroniciadc instead of 3-formyl-4-meth-
oxyphenylboronicacid. Yield 58%; mp 182–183 ꢁC. H
NMR (DMSO-d6); d 7.77 (t, J = 7.8Hz, 1H), 8.04 (d,
J = 7.8Hz, 1H), 8.29 (d, J = 8.4Hz, 1H), 8.43 (dd, J =
8.4, 2.1Hz, 1H), 8.48 (d, J = 7.8Hz, 1H), 8.69 (s, 1H),
9.13 (d, J = 2.1Hz, 1H), 10.13 (s, 1H). 13C NMR
(DMSO-d6); d 192.8, 157.7, 152.4, 141.1, 137.6, 136.8,
132.7, 130.7, 129.8, 128.3, 120.4, 116.9, 107.9. MS
1

M. A. Ismail et al. / Bioorg. Med. Chem. 12 (2004) 5405–5413
5413
7. Boykin, D. W.; Kumar, A.; Xiao, G.; Wilson, W. D.;
Bender, B. C.; McCurdy, D. R.; Hall, J. E.; Tidwell, R. R.
J. Med. Chem. 1998, 41, 124.
8. Francesconi, I.; Wilson, W. D.; Tanious, F. A., et al.
J. Med. Chem. 1999, 42, 2260.
9. Dykstra, C. C.; McClernon, D. R.; Elwell, L. P.; Tidwell,
R. R. Antimicrob. Agents Chemother. 1994, 38, 1890.
10. Bailly, C.; Dassonneville, L.; Carrascol, C.; Lucasl, D.;
Kumar, A.; Boykin, D. W.; Wilson, W. D. Anti-Cancer
Drug Des. 1999, 14, 47.
16. Del Poeta, M.; Schell, W. A.; Dykstra, C. C.; Jones, S.
K.; Tidwell, R. R.; Kumar, A.; Boykin, D. W.
Perfect, J. R. Antimicrob. Agents Chemother. 1998, 42,
2503.
17. Hopkins, K. T.; Wilson, W. D.; Bender, B. C.; McCurdy,
D. R.; Hall, J. E.; Tidwell, R. R.; Kumar, A.; Bajic, M.;
Boykin, D. W. J. Med. Chem. 1998, 41, 3872.
18. Wang, L.; Bailly, C.; Kumar, A.; Ding, D.; Bajic, M.;
Boykin, D. W.; Wilson, W. D. Proc. Natl. Acad. Sci.
U.S.A. 2000, 97, 12.
11. Fitzgerald, D. J.; Anderson, J. N. J. Biol. Chem. 1999, 274,
27128.
12. Ismail, M. A.; Brun, R.; Easterbrook, J. D.; Tanious, F.
A.; Wilson, W. D.; Boykin, D. W. J. Med. Chem. 2003, 46,
4761.
13. Tidwell, R. R.; Geratz, J. D.; Dann, O.; Volz, G.; Zeh, D.;
Loewe, H. J. Med. Chem. 1978, 21, 613.
14. Lombardy, R. L.; Tanious, F. A.; Ramachandran, K.;
Tidwell, R. R.; Wilson, W. D. J. Med. Chem. 1996, 39,
1452.
19. Wang, L.; Carrasco, C.; Kumar, A.; Stephens, C. E.;
Bailly, C.; Boykin, D. W.; Wilson, W. D. Biochemistry
2001, 40, 2511.
20. Tanious, F. A.; Wilson, W. D.; Wang, L.; Kumar, A.;
Boykin, D. W.; Marty, C.; Baldeyrou, B.; Bailly, C.
Biochemistry 2003, 42, 13576.
21. Nguyen, B.; Tardy, C.; Bailly, C.; Houssier, C.; Kumar,
A.; Boykin, D. W.; Wilson, W. D. Biopolymer 2002, 63,
281.
22. Ra¨z, B. M.; Iten, M.; Grether-Buhler, Y.; Kaminsky, R.;
¨
Brun, R. Acta Trop. 1997, 68, 139.
15. Del Poeta, M.; Schell, W. A.; Dykstra, C. C.; Jones, S. K.;
Tidwell, R. R.; Czarny, A.; Bajic, M.; Bajic, M.; Kumar,
A.; Boykin, D.; Perfect, J. R. Antimicrob. Agents Chemo-
ther. 1998, 42, 2495.
23. Verner, E.; Katz, B. A.; Spencer, J. R., et al. J. Med.
Chem. 2001, 44, 2753.
24. Forrest, H. S.; Walker, J. J. Chem. Soc. 1948, 1939.